DESCRIPTION: The goal of this study is to develop an approach to study the rules of viral adaptation at the molecular level that may inform us about how to develop anti-viral therapies. There are 3 specific aims to this proposal. (1) To examine the population genetics of viral populations over time. The data generated related to this aim will allow three hypotheses to be addressed. Is the pattern of substitution indicative of a series of selective sweeps? Are most of the fitness improvements due to a few substitutions with major effect and most of the substitutions will have very little effect? As the population reaches equilibrium will the level of variation decay to one sequence? (2) To explore the implications of parallel and convergent evolution during viral adaptation. Will parallel changes occur most of the time at sites with major as opposed to minor fitness benefits? Will related viruses adapt to the same selection with the same changes? (3) To examine the fitness cost of viral adaptation to multiple agents. Is there a fitness cost to adapting to a new selective agent? Can responses to multiple agents be predicted from responses to single agents? Is adaptation dependent on the prior history of adaptation? The PI proposes to use phage chemostats and adapt replicate populations to 43.5C. Samples will be taken each 6 hours for 3 days, then each 12 hours for 7 days. 96 samples will be taken each time point and assayed for a set of substitutions by oligonucleotide screening of predetermined regions (the set chosen from a pilot genome sequencing efforts). Fitness will be assayed (as the increase in phage per hour after infection of bacteria). These data will be used to address specific aim #1 (population genetics of viral adaptations over time) Six replicates of the first experiment will be undertaken (3 at a separate institution). These data will allow the PI to address specific aim #2 (the implications of convergent evolution during phage adaptation). One isolate from each replicate will be sequenced to determine screening sites. A second phase of research involves selecting against multiple agents. A series of agents, such as alcohol's, salts, etc. will be used as the selective agents. Selection will take place on a number of different media, as needed. Phage will be evolved that are resistant to a large number of agents, both separately and in combination. These data will allow the PI to address specific aim #3 (the fitness costs of adaptation to multiple agents).